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Barley (Hordeum vulgare L.) Transformation Using Immature Embryos

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Agrobacterium Protocols

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1223))

Abstract

Barley is a major crop species, and also has become a genetic model for the small grain temperate cereals. A draft barley genome sequence has recently been completed, opening many opportunities for candidate gene isolation and functionality testing. Thanks to the development of customizable endonucleases, also site-directed genome modification recently became feasible for higher plants, which marks the beginning of a new era of genetic engineering. The development of improved binary vectors and hypervirulent Agrobacterium tumefaciens strains has raised the efficiency of genetic transformation in barley to a level where the technique has become relatively routine. The transformation method described here involves immature barley embryos cocultivated with Agrobacterium after removal of their embryo axis. Critical adjustments to the protocol have included the supplementation of the cocultivation medium with the polyphenolic signaling compound acetosyringone at comparatively high concentration and the use of cysteine to reduce the extent of cellular oxidation upon agroinfection. In addition, the use of liquid, rather than solid, cocultivation medium promotes the throughput of the method. The protocol has delivered well over 10,000 transgenic barley plants over the past 10 years. Routine transformation efficiency, calculated on the basis of the recovery of independent transgenics per 100 explants, has reached about 25 % in cultivar (cv.) “Golden Promise”. The protocol has proven effective for more than 20 barley cultivars, although some adjustments to the culture conditions have had to be made in some cases. The transformation efficiency of cv. “Golden Promise” remains higher than that of any other cultivar tested.

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Acknowledgement

We are grateful to the Leibniz Institute of Plant Genetics and Crop Plant Research (IPK) Gatersleben for continuously providing excellent working conditions for our research group.

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Authors and Affiliations

  1. Plant Reproductive Biology, Leibniz Institute of Plant Genetics and Crop Plant Research (IPK), Corrensstr. 3, 06466, Stadt Seeland, OT Gatersleben, Germany

    Cornelia Marthe, Jochen Kumlehn & Goetz Hensel

Authors
  1. Cornelia Marthe
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  2. Jochen Kumlehn
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  3. Goetz Hensel
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Correspondence to Goetz Hensel .

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Editors and Affiliations

  1. Plant Sciences Institute, and Department of Agronomy, Iowa State University, Center for Plant Transformation, Ames, Iowa, USA

    Kan Wang

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Marthe, C., Kumlehn, J., Hensel, G. (2015). Barley (Hordeum vulgare L.) Transformation Using Immature Embryos. In: Wang, K. (eds) Agrobacterium Protocols. Methods in Molecular Biology, vol 1223. Springer, New York, NY. https://doi.org/10.1007/978-1-4939-1695-5_6

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  • DOI: https://doi.org/10.1007/978-1-4939-1695-5_6

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  • Publisher Name: Springer, New York, NY

  • Print ISBN: 978-1-4939-1694-8

  • Online ISBN: 978-1-4939-1695-5

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